Few natural host-parasite interactions have been highly characterized to date, even though the molecular mechanisms that determine parasite success or failure offer critical insights into immunity. We have developed a model system based on a coevolved host-parasite pair to analyze the reciprocal interactions underlying immunity in Drosophila melanogaster and the attack arsenals of its parasitic wasps (Leptopilina spp.). This system offers the complexity and subtlety to dissect innate immune activation, silencing, and subversion. D. melanogaster genetics are tractable, fast, and low-impact. Innate immune functions are widely conserved from fly to human and our long-term goal is to provide translational results. The most unique venom products of Leptopilina spp. are immunosuppressive microstructures, referred to as VLPs. Cell-specific drug delivery and therapeutic immune modulation represent two possible outcomes our investigations of the function of these unique parasite-derived particles. The protein composition of VLPs is central to the attack success of these parasites of Drosophila. This work marks the first functional investigation into a singular VLP protein. We have selected a gene product that is expressed only by the most virulent of Leptopilina wasps, is highly abundant in the VLP proteome, and possesses a putative site for GTP binding and hydrolysis. When analyzed in a genome-wide genetic interaction screen, this GTPase causes synthetic growth repression associated with impaired retrograde transport. In Aim 1, we will confirm these genetic interaction results and then test them in the context of GTP/GDP-locked mutants, as well as mutations that negatively impact normal vesicular trafficking. In Aim 2, we will express this protein in sub-populations of Drosophila blood cells to examine its subcellular localization and protein-protein interactions. We will test its impact on NF-?B-dependent signaling in fly larvae. The Toll- NF-?B pathway underlies activation of both the cellular and humoral arms of Drosophila's innate immune system and we believe that this GTPase-based modulation of retrograde transport may suppress normal immune signaling. The strategies utilized in this work depend on a paired model system approach. We have obtained preliminary information on the cellular function and targeting of a novel protein via a powerful yeast screen that provides information on thousands of genetic interactions. This approach has allowed us to design appropriate and efficient experimental plans in Drosophila. Our aims will provide the PI with in-depth skills in yeast and Drosophila genetics, transgenics, high-throughput genome-wide genetic screens, immunohistochemistry, and fluorescence imaging. This work will be strongly supported by a team that includes experts in the field in an interdisciplinary environment that will provide high-level training to the PI in immunity, multiple model system, genetics, and advanced molecular biology techniques.